The present invention relates to a pharmaceutical composition which is useful in particular for treating disorders linked to an arterial hypertension.
Arterial hypertension is a disorder whose causes are still unknown. However, it is known that the central nervous system plays an important role in regulating the cardiovascular system by controlling both the activity of the autonomic sympathetic nervous system and of the baroreflex, as well as the release of hypophyseal hormones.
Similarly, clinical and experimental work suggests that the activity of the central nervous system and of the peripheral sympathetic nerves participates in the genesis of arterial hypertension.
It has also been shown that a renin/angiotensin system also exists in the central nervous system. It turns out that it controls cardiovascular functions and homeostasis of body fluids. All the components of the systemic renin/angiotensin system, including the precursors and enzymes required for angiotensin formation and degradation, as well as angiotensin receptors, have already been identified in the brain.
In the systemic renin/angiotensin system, it is known in particular that angiotensin II is generated under the action of an essentially membrane-bound enzyme which belongs to the zinc metalloprotease group. This ectopeptidase is known under the name of angiotensin converting enzyme (ACE) since it transforms the inactive peptide angiotensin I into angiotensin II.
It turns out that this angiotensin II is transformed in vivo into angiotensin III (AngIII) under the action of another zinc ectopeptidase, which has recently been cloned, aminopeptidase A (APA), which removes the N-terminal aspartyl residue of angiotensin II to result in angiotensin III. This angiotensin III is itself destroyed by various peptidases including in particular aminopeptidase N (APN).
These two zinc ectopeptidases, APA and APN, belong to the thermolysin-type enzyme group and have significant homology between their amino acid sequences.
It has been demonstrated that the inhibition of the enzyme for converting angiotensin I into angiotensin II, ACE, present in the systemic renin-angiotensin system leads, via a blockage of angiotensin II formation, to a drop in arterial pressure which is particularly sensitive in individuals suffering from hypertension. It turns out that these inhibitors block peripheral ACE both in the circulation and especially in many tissues: vascular endothelium, lung, kidney, etc.
Consequently, up until now it has been suggested that angiotensin II is the principal mediator of the cerebral renin-angiotensin system, by analogy with the peripheral system.
In fact, contrary to what has been accepted, it seems that in the cerebral renin/angiotensin system, the critical step is not thought to be the formation of angiotensin II by action of ACE on angiotensin I, but the formation of angiotensin III by action of APA on angiotensin II. This determinant role of angiotensin III in the cerebral renin/angiotensin system is in particular reinforced by the results which are shown in Example 4 below.
This set of data thus tends to identify angiotensin III as the effector peptide of the cerebral renin/angiotensin system, it being responsible for the increase in arterial pressure. More specifically, in the brain, it appears that Ang III exerts a tonic stimulatory effect on the central control of arterial pressure.
The present invention is based precisely on the demonstration that angiotensin III plays an essential role in controlling arterial pressure at the central level.
More particularly, the present invention is directed towards providing a pharmaceutical composition which makes it possible to decrease arterial pressure and thus to oppose an angiotensin III-induced increase in pressure.
More specifically, it claims a pharmaceutical composition which is useful for decreasing arterial pressure, characterized in that it comprises, as an active principle, at least one selective aminopeptidase A inhibitor.
As mentioned previously, aminopeptidase A (APA) is an ectoenzyme which belongs to the zinc metalloprotease family, the bacterial model of which is thermolysin.
APA is a glycoprotein which is in the form of a homodimer.
The cloning of its cDNA has revealed that each monomer is composed of an anchoring domain which separates a short N-terminal cytosolic segment from a large C-terminal extracellular domain which contains the active site including more particularly the zinc-binding site.
Now, it turns out that APA exhibits 34% amino acid sequence identity with APN, which itself is involved in the degradation of angiotensin III. This homology is, moreover, the highest at precisely the active site present in the glycosylated extracellular domain.
It is clear that this sequence homology between APA and APN constitutes a handicap for obtaining inhibitors which are specific and selective with regard to APA.
As mentioned above, the inhibitor used in the composition claimed is an inhibitor which is selective with regard to APA. This selectivity is revealed in particular by an affinity which is multiplied approximately by at least a factor of 100 for APA compared to APN.
More specifically, a molecule which is considered according to the invention to be an inhibitor which is selective with regard to APA, is a molecule which satisfies at least one of the following criteria:
its inhibitory potency on APA in vitro is less than or equal to 10xe2x88x927 M,
it exhibits a selectivity factor of approximately 100 with respect to the enzymes aminopeptidase N, aminopeptidase B (EC 3.4.11.6) and neutral endopeptidase (EC 3.4.24.11), and the angiotensin converting enzyme (EC 3.4.15.1).
when injected in vivo via the intracerebroventricular or systemic (if it crosses the blood-brain barrier) route, it blocks the formation of angiotensin III.
As an APA inhibitor which most particularly suits the invention, mention may be made in particular of (S)-3-amino-4-mercaptobutylsulphonic acid or a salt thereof with a pharmaceutically acceptable acid or base.
As emerges from the examples presented below, sodium (S)-3-amino-4-mercaptobutylsulphonate, hereafter referred to under the symbol EC33, exhibits significant inhibitory activity with regard to APA.
This inhibitor advantageously exhibits a selectivity factor of 100 with respect to aminopeptidase N.
When injected in mice via the intracerebroventricular route, in an amount of 30 xcexcg, a significant increase is noted in the half-life of angiotensin II (30 xcexcg), of a factor of about 2.6 compared to that observed in a control animal. In parallel, it totally blocks the formation of angiotensin III in the hypothalamus.
Similarly, experiments carried out in normotensive (WKY) or hypertensive (SHR) rats show that the injection of EC33 makes it possible to significantly decrease arterial pressure. The hypertensive effects of this APA inhibitor, EC33, is at a maximum for a dose of 100 xcexcg. It is xe2x88x9222 mmHg in normotensive rats and xe2x88x9228 mmHg in hypertensive rats. The duration of action at this dose is on average between 40 and 60 minutes.
These results thus show that APA, the enzyme responsible for producing angiotensin III in the central nervous system, constitutes a novel therapeutic target of the cerebral renin/angiotensin system, and that the use of an APA inhibitor makes it possible to significantly reduce arterial pressure.
The pharmaceutical compositions claimed can optionally contain one or more pharmaceutically acceptable vehicles. These vehicles are chosen so as to constitute a pharmaceutical composition which can be administered conventionally via the oral, transmucous, parenteral or rectal route.
Their optimal methods of administration, doses and pharmaceutical forms can be determined according to the criteria generally taken into account in establishing a therapeutic treatment which is suited to a patient, such as for example the age or body weight of the patient, the seriousness of his general condition, the tolerance to the treatment and the observed side effects, etc.
Consequently, the compositions according to the invention are particularly advantageous for treating essential arterial hypertension during which the sympathetic hyperactivity often observed during the early phase is probably mediated by an increased activity of the cerebral renin/angiotensin system.
As non-limiting illustrations of the disorders which can be treated with the compositions claimed, mention may be made in particular of heart and kidney failures, hydrodynamic homeostasis disorders and proteinurea decrease in diabetics.
In addition, the compositions according to the invention can be advantageously used in addition to the blockers of the systemic renin/angiotensin system.
As representatives of these blockers, mention may be made in particular of converting enzyme inhibitors such as Enaprilate and angiotensin II receptor antagonists such as Losartan.
According to one variant of the invention, the composition claimed also comprises an angiotensin I converting enzyme inhibitor or an AT1 receptor antagonist.
This type of blocker of the systemic renin/angiotensin system proves to be effective in pathologies such as hypertension, congestive heart failure and left ventricular dysfunction after myocardial infarction, but also in improving proteinurea in diabetics and in reducing the progression of chronic kidney failure.
The compositions claimed will also be effective for treating the infections mentioned above, by also acting at the level of the central control of arterial pressure and by blocking the activity of the central renin/angiotensin system via their APA inhibitor.
The invention also extends to the use of an APA inhibitor as defined above combined, where appropriate, with a pharmaceutically acceptable vehicle for manufacturing a medicine which is useful for decreasing arterial pressure.
It can be in particular (S)-3-amino-4-mercaptobutylsulphonic acid or a pharmaceutically acceptable salt thereof with an acid or base.
FIG. 1: Evolution with time of the tritiated AngII content in murine hypothalamus in the presence or absence of EC33.
FIG. 2: Semi-logarithmic representation of the tritiated AngII contents in murine hypothalamus in the absence (control) or presence of EC33 (30 xcexcg).
FIG. 3: Effect of various doses of EC33 injected via the intracerebroventricular route on arterial pressure in SHR and WKY rats.
FIG. 4: Effect of various doses of EC33 in terms of duration of effect on arterial pressure in SHR and WKY rats.
FIG. 5: Effect on pressure of an APN inhibitor, PC18, in SHR rats.
FIG. 6: Effect in terms of duration of an APN inhibitor, PC18, on arterial pressure in SHR rats.
FIG. 7: Effect on pressure of an APN inhibitor, PC18, in normotensive rats in the presence of an antagonist of angiotensinergic receptors of type 1, AT1.
FIG. 8: Effect on arterial pressure of the inhibitor EC33 injected via the intracerebroventricular (i.c.v.) route or via the intravenous (i.v.) route in normotensive (WKY) or spontaneously hypertensive (SHR) conscious rats.